Inhibition of Gpx4-mediated ferroptosis alleviates cisplatin-induced hearing loss in C57BL/6 mice.

Cisplatin-induced hearing loss is a common side effect of cancer chemotherapy in clinics; however, the mechanism of cisplatin-induced ototoxicity is still not completely clarified. Cisplatin-induced ototoxicity is mainly associated with the production of reactive oxygen species, activation of apoptosis, and accumulation of intracellular lipid peroxidation, which also is involved in ferroptosis induction. In this study, the expression of TfR1, a ferroptosis biomarker, was upregulated in the outer hair cells of cisplatin-treated mice. Moreover, several key ferroptosis regulator genes were altered in cisplatin-damaged cochlear explants based on RNA sequencing, implying the induction of ferroptosis. Ferroptosis-related Gpx4 and Fsp1 knockout mice were established to investigate the specific mechanisms associated with ferroptosis in cochleae. Severe outer hair cell loss and progressive damage of synapses in inner hair cells were observed in Atoh1-Gpx4-/- mice. However, Fsp1-/- mice showed no significant hearing phenotype, demonstrating that Gpx4, but not Fsp1, may play an important role in the functional maintenance of HCs. Moreover, findings showed that FDA-approved luteolin could specifically inhibit ferroptosis and alleviate cisplatin-induced ototoxicity through decreased expression of transferrin and intracellular concentration of ferrous ions. This study indicated that ferroptosis inhibition through the reduction of intracellular ferrous ions might be a potential strategy to prevent cisplatin-induced hearing loss.

MAEL in human cancers and implications in prognostication and predicting benefit from immunotherapy over VEGFR/mTOR inhibitors in clear cell renal cell carcinoma: a bioinformatic analysis.

Maelstrom (MAEL), a novel cancer/testis-associated gene, may facilitate the initiation and progression of human malignancies, warranting comprehensive investigations. Single-cell and tissue-bulk transcriptomic data demonstrated higher MAEL expression in testis (spermatogonia/spermatocyte), kidney (proximal tubular cell), and brain (neuron/astrocyte), and corresponding cancers, including testicular germ cell tumor, glioma, papillary renal cell carcinoma, and clear cell renal cell carcinoma (ccRCC). Of these cancers, only in ccRCC did MAEL expression exhibit associations with both recurrence-free survival and overall survival. High MAEL expression was associated with an anti-inflammatory tumor immune microenvironment and VEGFR/mTOR activation in ccRCC tissues and high sensitivities to VEGFR/PI3K-AKT-mTOR inhibitors in ccRCC cell lines. Consistent with these, low rather than high MAEL expression indicated remarkable progression-free survival benefits from immune checkpoint inhibitor (ICI)-based immunotherapies over VEGFR/mTOR inhibitors in two large phase III trials (JAVELIN Renal 101 and CheckMate-025). MAEL is a biologically and clinically significant determinant with potential for prognostication after nephrectomy and patient selection for VEGFR/mTOR inhibitors and immunotherapy-based treatments.

Peroxisome Deficiency in Cochlear Hair Cells Causes Hearing Loss by Deregulating BK Channels.

The peroxisome is a ubiquitous organelle in rodent cells and plays important roles in a variety of cell types and tissues. It is previously indicated that peroxisomes are associated with auditory function, and patients with peroxisome biogenesis disorders (PBDs) are found to have hearing dysfunction, but the specific role of peroxisomes in hearing remains unclear. In this study, two peroxisome-deficient mouse models (Atoh1-Pex5-/- and Pax2-Pex5-/- ) are established and it is found that peroxisomes mainly function in the hair cells of cochleae. Furthermore, peroxisome deficiency-mediated negative effects on hearing do not involve mitochondrial dysfunction and oxidative damage. Although the mammalian target of rapamycin complex 1 (mTORC1) signaling is shown to function through peroxisomes, no changes are observed in the mTORC1 signaling in Atoh1-Pex5-/- mice when compared to wild-type (WT) mice. However, the expression of large-conductance, voltage-, and Ca2+ -activated K+ (BK) channels is less in Atoh1-Pex5-/- mice as compared to the WT mice, and the administration of activators of BK channels (NS-1619 and NS-11021) restores the auditory function in knockout mice. These results suggest that peroxisomes play an essential role in cochlear hair cells by regulating BK channels. Hence, BK channels appear as the probable target for treating peroxisome-related hearing diseases such as PBDs.

Dyslexia-Related Hearing Loss Occurs Mainly through the Abnormal Spontaneous Electrical Activity of Spiral Ganglion Neurons.

Dyslexia is a reading and spelling disorder due to neurodevelopmental abnormalities and is occasionally found to be accompanied by hearing loss, but the reason for the associated deafness remains unclear. This study finds that knockout of the dyslexia susceptibility 1 candidate 1 gene (Dyx1c1-/- ) in mice, the best gene for studying dyslexia, causes severe hearing loss, and thus it is a good model for studying the mechanism of dyslexia-related hearing loss (DRHL). This work finds that the Dyx1c1 gene is highly expressed in the mouse cochlea and that the spontaneous electrical activity of inner hair cells and type I spiral ganglion neurons is altered in the cochleae of Dyx1c1-/- mice. In addition, primary ciliary dyskinesia-related phenotypes such as situs inversus and disrupted ciliary structure are seen in Dyx1c1-/- mice. In conclusion, this study gives new insights into the mechanism of DRHL in detail and suggests that Dyx1c1 may serve as a potential target for the clinical diagnosis of DRHL.

Growth of spiral ganglion neurons induced by graphene oxide/oxidized bacterial cellulose composite hydrogel.

The damage or degeneration of spiral ganglion neurons (SGNs) can impair the auditory signals transduction from hair cells to the central auditory system, and cause significant hearing loss. Herein, a new form of bioactive hydrogel incorporating topological graphene oxide (GO) and TEMPO-oxidized bacterial cellulose (GO/TOBC hydrogel) was developed to provide a favorable microenvironment for SGN neurite outgrowth. As the network structure of lamellar interspersed fiber cross-linked by GO/TOBC hydrogels well simulated the structure and morphology of ECM, with the controllable hydrophilic property and appropriate Young's modulus well met those requirements of SGNs microenvironment, the GO/TOBC hybrid matrix exhibited great potential to promote the growth of SGNs. The quantitative real-time PCR result confirmed that the GO/TOBC hydrogel can significantly accelerate the development of growth cones and filopodia, by increasing the mRNA expression levels of diap3, fscn2, and integrin ß1. These results suggest that GO/TOBC hydrogel scaffolds have the potential to be used to construct biomimetic nerve grafts for repairing or replacing nerve defects.

Macrophage-mediated immune response aggravates hearing disfunction caused by the disorder of mitochondrial dynamics in cochlear hair cells.

Mitochondrial dynamics is essential for maintaining the physiological function of the mitochondrial network, and its disorders lead to a variety of diseases. Our previous study identified mitochondrial dynamics controlled anti-tumor immune responses and anxiety symptoms. However, how mitochondrial dynamics affects auditory function in the inner ear remains unclear. Here, we show that the deficiency of FAM73a or FAM73b, two mitochondrial outer membrane proteins that mediate mitochondrial fusion, leads to outer hair cells (HCs) damage and progressive hearing loss in FVB/N mice. Abnormal mitochondrial fusion causes elevated oxidative stress and apoptosis of HCs in the early stage. Thereafter, the activation of macrophages and CD4+ T cell is found in the mutant mice with the increased expression of the inflammatory cytokines IL-12 and IFN-γ compared with control mice. Strikingly, a dramatically decreased number of macrophages by Clophosome®-A-Clodronate Liposomes treatment alleviates the hearing loss of mutant mice. Collectively, our finding highlights that FAM73a or FAM73b deficiency affects HCs survival by disturbing the mitochondrial function, and the subsequent immune response in the cochleae worsens the damage of HCs.

Gstm1/Gstt1 is essential for reducing cisplatin ototoxicity in CBA/CaJ mice.

Cisplatin is a widely used chemotherapeutic agent. However, its clinical utility is limited because of cisplatin-induced ototoxicity. Glutathione S-transferase (GST) was found to play a vital role in reducing cisplatin ototoxicity in mice. Deletion polymorphisms of GSTM1 and GSTT1, members of the GST family, are common in humans and are presumed to be associated with cisplatin-induced hearing impairment. However, the specific roles of GSTM1 and GSTT1 in cisplatin ototoxicity are not completely clear. Here, under cisplatin treatment, simultaneous deletion of Gstm1 and Gstt1 lead to a more profound hearing loss in CBA/CaJ mice (Gstm1/Gstt1-DKO) than in wild-type mice. The Gstm1/Gstt1-DKO mice, in which phase II detoxification genes were upregulated, exhibited more severe oxidative stress and higher outer hair cell apoptosis in the cochleae than the control mice. Thus, our study revealed that Gstm1 and Gstt1 protect auditory hair cells from cisplatin-induced ototoxicity in the CBA/CaJ mice, and genetic screening for GSTM1 and GSTT1 polymorphisms could help determine a standard cisplatin dose for cancer patients undergoing chemotherapy.

Developing Strategy to Predict the Results of Prostate Multiparametric Magnetic Resonance Imaging and Reduce Unnecessary Multiparametric Magnetic Resonance Imaging Scan.

PURPOSE: The clinical utility of multiparametric magnetic resonance imaging (mpMRI) for the detection and localization of prostate cancer (PCa) has been evaluated and validated. However, the implementation of mpMRI into the clinical practice remains some burden of cost and availability for patients and society. We aimed to predict the results of prostate mpMRI using the clinical parameters and multivariable model to reduce unnecessary mpMRI scans. METHODS: We retrospectively identified 784 men who underwent mpMRI scans and subsequent prostate biopsy between 2016 and 2020 according to the inclusion criterion. The cohort was split into a training cohort of 548 (70%) patients and a validation cohort of 236 (30%) patients. Clinical parameters including age, prostate-specific antigen (PSA) derivates, and prostate volume (PV) were assessed as the predictors of mpMRI results. The mpMRI results were divided into groups according to the reports: "negative", "equivocal", and "suspicious" for the presence of PCa. RESULTS: Univariate analysis showed that the total PSA (tPSA), free PSA (fPSA), PV, and PSA density (PSAD) were significant predictors for suspicious mpMRI (P < 0.05). The PSAD (AUC = 0.77) and tPSA (AUC = 0.74) outperformed fPSA (AUC = 0.68) and PV (AUC = 0.62) in the prediction of the mpMRI results. The multivariate model (AUC = 0.80) had a similar diagnostic accuracy with PSAD (P = 0.108), while higher than tPSA (P = 0.024) in predicting the mpMRI results. The multivariate model illustrated a better calibration and substantial improvement in the decision curve analysis (DCA) at a threshold above 20%. Using the PSAD with a 0.13 ng/ml2 cut-off could spare the number of mpMRI scans by 20%, keeping a 90% sensitivity in the prediction of suspicious MRI-PCa and missing three (3/73, 4%) clinically significant PCa cases. At the same sensitivity level, the multivariate model with a 32% cut-off could spare the number of mpMRI scans by 27%, missing only one (1/73, 1%) clinically significant PCa case. CONCLUSION: Our multivariate model could reduce the number of unnecessary mpMRI scans without comprising the diagnostic ability of clinically significant PCa. Further prospective validation is required.

Morphological diversity of single neurons in molecularly defined cell types.

Dendritic and axonal morphology reflects the input and output of neurons and is a defining feature of neuronal types1,2, yet our knowledge of its diversity remains limited. Here, to systematically examine complete single-neuron morphologies on a brain-wide scale, we established a pipeline encompassing sparse labelling, whole-brain imaging, reconstruction, registration and analysis. We fully reconstructed 1,741 neurons from cortex, claustrum, thalamus, striatum and other brain regions in mice. We identified 11 major projection neuron types with distinct morphological features and corresponding transcriptomic identities. Extensive projectional diversity was found within each of these major types, on the basis of which some types were clustered into more refined subtypes. This diversity follows a set of generalizable principles that govern long-range axonal projections at different levels, including molecular correspondence, divergent or convergent projection, axon termination pattern, regional specificity, topography, and individual cell variability. Although clear concordance with transcriptomic profiles is evident at the level of major projection type, fine-grained morphological diversity often does not readily correlate with transcriptomic subtypes derived from unsupervised clustering, highlighting the need for single-cell cross-modality studies. Overall, our study demonstrates the crucial need for quantitative description of complete single-cell anatomy in cell-type classification, as single-cell morphological diversity reveals a plethora of ways in which different cell types and their individual members may contribute to the configuration and function of their respective circuits.

Development and head-to-head comparison of machine-learning models to identify patients requiring prostate biopsy.

BACKGROUND: Machine learning has many attractive theoretic properties, specifically, the ability to handle non predefined relations. Additionally, studies have validated the clinical utility of mpMRI for the detection and localization of CSPCa (Gleason score ≥ 3 + 4). In this study, we sought to develop and compare machine-learning models incorporating mpMRI parameters with traditional logistic regression analysis for prediction of PCa (Gleason score ≥ 3 + 3) and CSPCa on initial biopsy. METHODS: A total of 688 patients with no prior prostate cancer diagnosis and tPSA ≤ 50 ng/ml, who underwent mpMRI and prostate biopsy were included between 2016 and 2020. We used four supervised machine-learning algorithms in a hypothesis-free manner to build models to predict PCa and CSPCa. The machine-learning models were compared to the logistic regression analysis using AUC, calibration plot, and decision curve analysis. RESULTS: The artificial neural network (ANN), support vector machine (SVM), and random forest (RF) yielded similar diagnostic accuracy with logistic regression, while classification and regression tree (CART, AUC = 0.834 and 0.867) had significantly lower diagnostic accuracy than logistic regression (AUC = 0.894 and 0.917) in prediction of PCa and CSPCa (all P < 0.05). However, the CART illustrated best calibration for PCa (SSR = 0.027) and CSPCa (SSR = 0.033). The ANN, SVM, RF, and LR for PCa had higher net benefit than CART across the threshold probabilities above 5%, and the five models for CSPCa displayed similar net benefit across the threshold probabilities below 40%. The RF (53% and 57%, respectively) and SVM (52% and 55%, respectively) for PCa and CSPCa spared more unnecessary biopsies than logistic regression (35% and 47%, respectively) at 95% sensitivity for detection of CSPCa. CONCLUSION: Machine-learning models (SVM and RF) yielded similar diagnostic accuracy and net benefit, while spared more biopsies at 95% sensitivity for detection of CSPCa, compared with logistic regression. However, no method achieved desired performance. All methods should continue to be explored and used in complementary ways.

Deletion of Kcnj16 in Mice Does Not Alter Auditory Function.

Endolymphatic potential (EP) is the main driving force behind the sensory transduction of hearing, and K+ is the main charge carrier. Kir5.1 is a K+ transporter that plays a significant role in maintaining EP homeostasis, but the expression pattern and role of Kir5.1 (which is encoded by the Kcnj16 gene) in the mouse auditory system has remained unclear. In this study, we found that Kir5.1 was expressed in the mouse cochlea. We checked the inner ear morphology and measured auditory function in Kcnj16 -/- mice and found that loss of Kcnj16 did not appear to affect the development of hair cells. There was no significant difference in auditory function between Kcnj16 -/- mice and wild-type littermates, although the expression of Kcnma1, Kcnq4, and Kcne1 were significantly decreased in the Kcnj16 -/- mice. Additionally, no significant differences were found in the number or distribution of ribbon synapses between the Kcnj16 -/- and wild-type mice. In summary, our results suggest that the Kcnj16 gene is not essential for auditory function in mice.

Multivariable Models Incorporating Multiparametric Magnetic Resonance Imaging Efficiently Predict Results of Prostate Biopsy and Reduce Unnecessary Biopsy.

PURPOSE: We sought to develop diagnostic models incorporating mpMRI examination to identify PCa (Gleason score≥3+3) and CSPCa (Gleason score≥3+4) to reduce overdiagnosis and overtreatment. METHODS: We retrospectively identified 784 patients according to inclusion criteria between 2016 and 2020. The cohort was split into a training cohort of 548 (70%) patients and a validation cohort of 236 (30%) patients. Age, PSA derivatives, prostate volume, and mpMRI parameters were assessed as predictors for PCa and CSPCa. The multivariable models based on clinical parameters were evaluated using area under the curve (AUC), calibration plots, and decision curve analysis (DCA). RESULTS: Univariate analysis showed that age, tPSA, PSAD, prostate volume, MRI-PCa, MRI-seminal vesicle invasion, and MRI-lymph node invasion were significant predictors for both PCa and CSPCa (each p≤0.001). PSAD has the highest diagnostic accuracy in predicting PCa (AUC=0.79) and CSPCa (AUC=0.79). The multivariable models for PCa (AUC=0.92, 95% CI: 0.88-0.96) and CSPCa (AUC=0.95, 95% CI: 0.92-0.97) were significantly higher than the combination of derivatives for PSA (p=0.041 and 0.009 for PCa and CSPCa, respectively) or mpMRI (each p<0.001) in diagnostic accuracy. And the multivariable models for PCa and CSPCa illustrated better calibration and substantial improvement in DCA at threshold above 10%, compared with PSA or mpMRI derivatives. The PCa model with a 30% cutoff or CSPCa model with a 20% cutoff could spare the number of biopsies by 53%, and avoid the number of benign biopsies over 80%, while keeping a 95% sensitivity for detecting CSPCa. CONCLUSION: Our multivariable models could reduce unnecessary biopsy without comprising the ability to diagnose CSPCa. Further prospective validation is required.

Knockdown of Foxg1 in supporting cells increases the trans-differentiation of supporting cells into hair cells in the neonatal mouse cochlea.

Foxg1 is one of the forkhead box genes that are involved in morphogenesis, cell fate determination, and proliferation, and Foxg1 was previously reported to be required for morphogenesis of the mammalian inner ear. However, Foxg1 knock-out mice die at birth, and thus the role of Foxg1 in regulating hair cell (HC) regeneration after birth remains unclear. Here we used Sox2CreER/+ Foxg1loxp/loxp mice and Lgr5-EGFPCreER/+ Foxg1loxp/loxp mice to conditionally knock down Foxg1 specifically in Sox2+ SCs and Lgr5+ progenitors, respectively, in neonatal mice. We found that Foxg1 conditional knockdown (cKD) in Sox2+ SCs and Lgr5+ progenitors at postnatal day (P)1 both led to large numbers of extra HCs, especially extra inner HCs (IHCs) at P7, and these extra IHCs with normal hair bundles and synapses could survive at least to P30. The EdU assay failed to detect any EdU+ SCs, while the SC number was significantly decreased in Foxg1 cKD mice, and lineage tracing data showed that much more tdTomato+ HCs originated from Sox2+ SCs in Foxg1 cKD mice compared to the control mice. Moreover, the sphere-forming assay showed that Foxg1 cKD in Lgr5+ progenitors did not significantly change their sphere-forming ability. All these results suggest that Foxg1 cKD promotes HC regeneration and leads to large numbers of extra HCs probably by inducing direct trans-differentiation of SCs and progenitors to HCs. Real-time qPCR showed that cell cycle and Notch signaling pathways were significantly down-regulated in Foxg1 cKD mice cochlear SCs. Together, this study provides new evidence for the role of Foxg1 in regulating HC regeneration from SCs and progenitors in the neonatal mouse cochlea.

A miniature mass spectrometer for liquid chromatography applications.

A miniature mass spectrometer capable of detecting analytes eluting from a high-performance liquid chromatography (HPLC) system is described and demonstrated for the first time. The entire instrument, including all pumps and the computer, is contained within a single enclosure that may be conveniently accommodated at the base of the HPLC stack. The microspray ion source, vacuum interface, ion guide, and quadrupole ion filter are all microengineered. These components are fabricated in batches using microelectromechanical systems (MEMS) techniques and considered to be consumables. When coupled to a standard HPLC system using an integrated passive split, the limit of detection for reserpine while scanning the full mass range is 5 ng on-column (1 pg of which is passed to the microspray). The mass range is m/z 100-800, and each spectrum is typically acquired at a rate of 1 scan per second.

Comparison of ion coupling strategies for a microengineered quadrupole mass filter.

The limitations of conventional machining and assembly techniques require that designs for quadrupole mass analyzers with rod diameters less than a millimeter are not merely scale versions of larger instruments. We show how silicon planar processing techniques and microelectromechanical systems (MEMS) design concepts can be used to incorporate complex features into the construction of a miniature quadrupole mass filter chip that could not easily be achieved using other microengineering approaches. Three designs for the entrance and exit to the filter consistent with the chosen materials and techniques have been evaluated. The differences between these seemingly similar structures have a significant effect on the performance. Although one of the designs results in severe attenuation of transmission with increasing mass, the other two can be scanned to m/z = 400 without any corruption of the mass spectrum. At m/z = 219, the variation in the transmission of the three designs was found to be approximately four orders of magnitude. A maximum resolution of M/DeltaM = 87 at 10% peak height has been achieved at m/z = 219 with a filter operated at 6 MHz and constructed using rods measuring (508 +/- 5) microm in diameter.